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Structural basis of cell surface receptor recognition by botulinum neurotoxin B

Abstract

Botulinum neurotoxins (BoNTs) are potent bacterial toxins that cause paralysis at femtomolar concentrations1 by blocking neurotransmitter release. A ‘double receptor’ model has been proposed in which BoNTs recognize nerve terminals via interactions with both gangliosides and protein receptors that mediate their entry2. Of seven BoNTs (subtypes A–G), the putative receptors for BoNT/A3,4, BoNT/B5,6 and BoNT/G7 have been identified, but the molecular details that govern recognition remain undefined. Here we report the crystal structure of full-length BoNT/B in complex with the synaptotagmin II (Syt-II) recognition domain at 2.6 Å resolution. The structure of the complex reveals that Syt-II forms a short helix that binds to a hydrophobic groove within the binding domain of BoNT/B. In addition, mutagenesis of amino acid residues within this interface on Syt-II affects binding of BoNT/B. Structural and sequence analysis reveals that this hydrophobic groove is conserved in the BoNT/G and BoNT/B subtypes, but varies in other clostridial neurotoxins. Furthermore, molecular docking studies using the ganglioside GT1b indicate that its binding site is more extensive than previously proposed and might form contacts with both BoNT/B and synaptotagmin. The results provide structural insights into how BoNTs recognize protein receptors and reveal a promising target for blocking toxin–receptor recognition.

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Figure 1: The structure of BoNT/B–Syt-II recognition domain complex.
Figure 2: Interaction between the H CC domain of BoNT/B and the recognition domain of Syt-II.
Figure 3: Ganglioside-supported BoNT/B binding with synaptotagmin.
Figure 4: Model of BoNT/B binding to its receptors on a neuron.

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Acknowledgements

We thank E. Abola, K. Saikatendu and J. Ng for discussions and A. Walker for assistance with manuscript preparation. This work was supported by a grant from the Pacific Southwest Regional Center of Excellence (R.C.S. and E.A.J.), and grants from the NIH/NIAID (to E.R.C.). E.R.C. and E.A.J. acknowledge membership of and support from the Region V “Great Lakes” Regional Center of Excellence in Biodefense and Emerging Infectious Diseases. E.R.C. is an Investigator of the Howard Hughes Medical Institute. Portions of this research were carried out at the SSRL, a national user facility operated by Stanford University on behalf of the US Department of Energy, Office of Basic Energy Sciences. The SSRL Structural Molecular Biology Program is supported by the Department of Energy, Office of Biological and Environmental Research and by the National Institutes of Health, National Center for Research Resources, Biomedical Technology Program, and the National Institute of General Medical Sciences.

Author Contributions J.W.A., Q.C. (crystallography) and M.D. (molecular biology) contributed equally to this work and co-wrote the paper. All authors discussed the results and commented upon the manuscript.

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Correspondence to Edwin R. Chapman or Raymond C. Stevens.

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Atomic coordinates and experimental structure factors of the BoNT/B–Syt II ectodomain complex have been deposited in the RCSB Protein Data Bank (PDB) under accession code 2NP0. Reprints and permissions information is available at www.nature.com/reprints. The authors declare no competing financial interests.

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This file contains Supplementary Methods, Supplementary Table 1 and 2 and Supplementary Figures 1-7 with legends.

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Chai, Q., Arndt, J., Dong, M. et al. Structural basis of cell surface receptor recognition by botulinum neurotoxin B . Nature 444, 1096–1100 (2006). https://doi.org/10.1038/nature05411

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